From 1990 through 1993 the Wadsworth Center Biological Microscopy and Image Reconstruction Resource (BMIRR) was used by more than 100 biomedical researchers, over half of which were funded by NIH. During this time these users published more than 100 full Technological Research and Development (TRD), Collaborative and Service papers, and also over 100 abstracts, acknowledging the contribution of the BMIRR. While the BMIRR remains popular with cell and neurobiologists, an in-creasing number of NIH-funded researchers working on biomaterials/implants, bone structure and formation, and development also found that the BMIRR substantially benefitted their research programs during the last grant cycle. The main features of the BMIRR include a 1.2 MV AEI HVEM, a 400 KV JEOL 4000 IVEM, confocal and video-enhanced light microscope (LM) facilities with laser trapping and microsurgery systems, and computer- based image processing for producing contour (STERECON) and tomographic 3-D ultrastructural reconstructions. These capabilities allow users to conduct correlative LM and EM studies at the highest possible resolution and to obtain 3-D ultrastructural data encompassing a wide range of cellular organization from macromolecules/organelles to whole cells/tissues. The objective of the proposed renewal is to further develop and expand BMIRR'S capabilities so that it will service an even greater number of NIH-funded research programs. The specific aims of the three TRD areas include: 1) To continue making this state-of-the-art microscopy and image reconstruction facility, and those methods and components developed under previous NCRR awards, available to the biomedical research community. This will include modernizing aspects of HVEM instrumentation and operation; 2) To develop and implement procedures for electron crystallography of proteins (especially membrane proteins) and small molecules such as lipids. This includes optimizing kinematic diffraction data collection over the voltage range 100 to 1200 KV, and developing phase-extension algorithms that utilize direct methods combined with high-resolution images; and 3) To improve existing procedures for tomographically reconstructing biological specimens from tilted IVEM/HVEM images, and make them applicable to frozen-hydrated specimens. This will require implementing automated data collection for low dose imaging as well as developing software for improved data alignment, reconstruction and restoration.